Crosslinked polymers with amine binding groups

ABSTRACT

Crosslinked polymeric materials are described that contain pendant amine capture groups. The amine capture groups include N-sulfonyldicarboximide groups that can react with amine-containing materials by a ring opening reaction. Reaction mixtures used to prepare the crosslinked polymeric materials, articles containing the crosslinked polymeric materials, methods of making articles, and methods of immobilizing an amine-containing material are also described.

TECHNICAL FIELD

Crosslinked polymeric materials having pendant amine capture groups,articles containing the crosslinked polymeric materials, and methods ofimmobilizing an amine-containing material are described.

BACKGROUND

Amine-containing materials such as amine-containing analytes, aminoacids, DNA, RNA, proteins, cells, tissue, organelles, immunoglobins, orfragments thereof immobilized on a surface can be used in numerousapplications. For example, immobilized biological amines can be used forthe medical diagnosis of a disease or genetic defect, for biologicalseparations, or for detection of various biomolecules. Immobilization ofthe amine containing material is typically accomplished by reaction ofan amino group with an amine reactive functional group that iscovalently attached to the surface of the substrate.

Such amine reactive surfaces can be prepared, for example, by coatingonto the surface of a substrate a solution of a polymeric materialprepared from an amine reactive monomer such asN-[(meth)acryloxy]succinimide or vinyl azlactone. An amine-containingmaterial can react with the N-acyloxysuccinimide group resulting indisplacement of N-hydroxysuccinimide and formation of a carboxamide. Anamine-containing material can react with the cyclic azlactone resultingin an opening of the ring structure.

Although polymeric surfaces that include a reactive functional groupsuch as an N-acyloxysuccinimide group or an azlactone group can reactreadily with primary or secondary amine-containing materials, suchreactive functional groups can suffer from a number of disadvantages.For example, many of the reactions with biological amines are conductedin dilute aqueous solutions. Under these conditions, theN-acyloxysuccinimide functional group is known to undergo rapidhydrolysis. This competing reaction can cause incomplete or inefficientimmobilization of the amine-containing materials on the substrate.

While azlactone functional groups are more stable to hydrolysis, it isdifficult to synthesize an azlactone linked to any polymerizable groupother than a vinyl group. This results in polymeric material with theamine reactive functional group directly attached to the polymerbackbone. This can make it difficult for the amine containing materialto get close enough to the amine reactive group for efficientimmobilization.

A need exists for polymeric materials with alternative amine reactivefunctional groups that can be used as coatings for the immobilization ofamine-containing materials and that have good adhesion to a substrate.

SUMMARY

Reaction mixtures and crosslinked polymeric materials formed from thereaction mixtures are described. More specifically, the reactionmixtures and the crosslinked polymeric materials contain amine capturegroups. The amine capture groups include N-sulfonyldicarboximide groupsthat can react with amine-containing materials by a ring openingreaction. Articles containing the crosslinked polymeric material,methods of making the articles, and methods of immobilizing anamine-containing material are also described.

In a first aspect, a reaction mixture is described that includes (a) anamine capture monomer of Formula I

and (b) a crosslinking monomer that includes at least two (meth)acryloylgroups. The amine capture monomer of Formula I can be unsubstituted orsubstituted with a halo, alkyl, alkoxy, or combinations thereof. InFormula I,

-   -   L is an oxy or —NR⁶—;    -   R¹ and R² together with a dicarboximide group to which they are        attached form a four to eight membered heterocyclic or        heterobicyclic group that can be fused to an optional aromatic        group, optional saturated or unsaturated cyclic group, or        optional saturated or unsaturated bicyclic group;    -   R³ is hydrogen or methyl;    -   R⁴ is an alkyl, aryl, aralkyl, or —N(R⁵)₂, wherein each R⁵ is an        alkyl group or both R⁵ groups taken together with the nitrogen        atom to which they are attached form a four to eight membered        heterocyclic group;    -   R⁶ is hydrogen, alkyl, aryl, aralkyl, acyl, alkylsulfonyl, or        arylsulfonyl; and    -   Y is a single bond or a divalent group comprising an alkylene,        heteroalkylene, arylene, or combinations thereof.

In a second aspect, a crosslinked polymeric material is described thatincludes the reaction product of a reaction mixture that contains (a) anamine capture monomer of Formula I and (b) a crosslinking monomer thatincludes at least two (meth)acryloyl groups.

In a third aspect, an article is described that includes a substrate anda crosslinked polymeric material disposed on a surface of the substrate.The crosslinked polymeric material includes the reaction product of areaction mixture that contains (a) an amine capture monomer of Formula Iand (b) a crosslinking monomer that includes at least two (meth)acryloylgroups. The crosslinked polymeric material has a pendant amine capturegroup that includes a N-sulfonyldicarboximide group.

In a fourth aspect, a method of making an article is described. Themethod includes providing a substrate, disposing a reaction mixture on asurface of the substrate, and curing the reaction mixture to form acrosslinked polymeric material. The reaction mixture contains (a) anamine capture monomer of Formula I and (b) a crosslinking monomer thatincludes at least two (meth)acryloyl groups. The crosslinked polymericmaterial has a pendant amine capture group that includes aN-sulfonyldicarboximide group.

In a fifth aspect, a method of immobilizing an amine-containing materialis described. The method includes providing a substrate, and disposing areaction mixture on a surface of the substrate. The reaction mixturecontains (a) an amine capture monomer of Formula I and (b) acrosslinking monomer that includes at least two (meth)acryloyl groups.The method further includes curing the reaction mixture to form acrosslinked polymeric material having a pendant amine capture group thatincludes a N-sulfonyidicarboximide group, and reacting anamine-containing material with the N-sulfonyidicarboximide group.

In a sixth aspect, a polymeric material is provided that includespendant groups of Formula II

wherein

-   -   L is an oxy or —NR⁶—;    -   R¹ and R² together with a dicarboximide group to which they are        attached form a four to eight membered heterocyclic or        heterobicyclic group that can be fused to an optional aromatic        group, optional saturated or unsaturated cyclic group, or        optional saturated or unsaturated bicyclic group;    -   R⁴ is an alkyl, aryl, aralkyl, or —N(R⁵)₂, wherein each R⁵ is an        alkyl group or both R⁵ groups taken together with the nitrogen        atom to which they are attached form a four to eight membered        heterocyclic group;    -   R⁶ is hydrogen, alkyl, aryl, aralkyl, acyl, alkylsulfonyl, or        arylsulfonyl;    -   Y is a single bond or a divalent group comprising an alkylene,        heteroalkylene, arylene, or combinations thereof;    -   an asterisk (*) denotes an attachment site of the pendant group        to a backbone of the polymeric material;    -   the pendant group of Formula II is unsubstituted or substituted        with a halo, alkyl, alkoxy, or combinations thereof; and    -   the polymeric material is crosslinked.

In a seventh aspect, a polymeric material is provided that includespendant groups of Formula III

wherein

-   -   L is an oxy or —NR⁶—;    -   R⁴ is an alkyl, aryl, aralkyl, or —N(R⁵)₂, wherein each R⁵ is an        alkyl group or both R⁵ groups taken together with the nitrogen        atom to which they are attached form a four to eight membered        heterocyclic group;    -   R⁶ is hydrogen, alkyl, aryl, aralkyl, acyl, alkylsulfonyl, or        arylsulfonyl;    -   R⁷ is an alkylene having 1 to 5 carbon atoms, aromatic group,        saturated or unsaturated cyclic group, saturated or unsaturated        bicyclic group, or combination thereof;    -   T is equal to a primary amine-containing biological material        minus a —NH₂ group;    -   Y is a single bond or a divalent group comprising an alkylene,        heteroalkylene, arylene, or combinations thereof;    -   an asterisk (*) denotes an attachment site of the pendant group        to a backbone of the polymeric material;    -   the pendant group of Formula II is unsubstituted or substituted        with a halo, alkyl, alkoxy, or combinations thereof, and    -   the polymeric material is crosslinked.

The terms “a”, “an”, and “the” are used interchangeably with “at leastone” to mean one or more of the elements being described.

The term “acyl” refers to a monovalent group of formula —(CO)R where Ris an alkyl group and where (CO) used herein indicates that the carbonis attached to the oxygen with a double bond.

The term “alkoxy” refers to a monovalent group of formula —OR where R isan alkyl group.

The term “alkyl” refers to a monovalent group that is a radical of analkane and includes groups that are linear, branched, cyclic, orcombinations thereof. The alkyl group typically has 1 to 30 carbonatoms. In some embodiments, the alkyl group contains 1 to 20 carbonatoms, 1 to 10 carbon atoms, 1 to 6 carbon atoms, or 1 to 4 carbonatoms. Examples of alkyl groups include, but are not limited to, methyl,ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, n-pentyl,n-hexyl, cyclohexyl, n-heptyl, n-octyl, and ethylhexyl.

The term “alkylene” refers to a divalent group that is a radical of analkane. The alkylene can be straight-chained, branched, cyclic, orcombinations thereof. The alkylene typically has 1 to 30 carbon atoms.In some embodiments, the alkylene contains 1 to 20, 1 to 10, 1 to 8, 1to 6, or 1 to 4 carbon atoms. The radical centers of the alkylene can beon the same carbon atom (i.e., an alkylidene) or on different carbonatoms.

The term “alkylsulfonyl” refers to a monovalent group of formula —SO₂Rwhere R is an alkyl.

The term “amine capture monomer” refers to a monomer having an aminecapture group. The term “amine capture” refers to a group on a monomeror polymeric material that is capable of reacting with anamine-containing material. The amine capture group often includes aN-sulfonyidicarboximide group.

The term “amine-containing material” refers to a material that has aprimary amine group, a secondary amine group, or a combination thereof.The amine-containing material can be a biological material or anon-biological material. The amine-containing material often has analkylene group attached to the primary amine group, secondary aminegroup, or a combination thereof.

The term “aralkyl” refers to a monovalent group that is a radical of thecompound R—Ar where Ar is an aromatic carbocyclic group and R is analkyl group.

The term “aryl” refers to a monovalent group that is aromatic andcarbocyclic. The aryl can have one to five rings that are connected toor fused to the aromatic ring. The other ring structures can bearomatic, non-aromatic, or combinations thereof. Examples of aryl groupsinclude, but are not limited to, phenyl, biphenyl, terphenyl, anthryl,naphthyl, acenaphthyl, anthraquinonyl, phenanthryl, anthracenyl,pyrenyl, perylenyl, and fluorenyl.

The term “arylsulfonyl” refers to a monovalent group of formula —SO₂Arwhere Ar is an aryl.

The term “arylene” refers to a divalent group that is carbocyclic andaromatic. The group has one to five rings that are connected, fused, orcombinations thereof The other rings can be aromatic, non-aromatic, orcombinations thereof. In some embodiments, the arylene group has up to 5rings, up to 4 rings, up to 3 rings, up to 2 rings, or one aromaticring. For example, the arylene group can be phenylene.

The term “carbonyl” refers to a divalent group of formula —(CO)—.

The term “carbonylimino” refers to a divalent group of formula—(CO)NR^(b)— where R^(b) is hydrogen, alkyl, aryl, aralkyl, acyl,arylsulfonyl, or alkylsulfonyl.

The term “carbonyloxy” refers to a divalent group of formula —(CO)O—.

The term “dicarboximide” refers to a trivalent group of formula

The term “halo” refers to fluoro, bromo, chloro, or iodo.

The term “heteroalkylene” refers to a divalent alkylene having one ormore —CH₂— groups replaced with a thio, oxy, or —NR^(a)— where R^(a) ishydrogen or alkyl. The heteroalkylene can be linear, branched, cyclic,or combinations thereof and can include up to 60 carbon atoms and up to15 heteroatoms. In some embodiments, the heteroalkylene includes up to50 carbon atoms, up to 40 carbon atoms, up to 30 carbon atoms, up to 20carbon atoms, or up to 10 carbon atoms.

The term “(meth)acrylate” refers to monomer that is an acrylate ormethacrylate. Likewise, the term “(meth)acrylamide” refers to a monomerthat is an acrylamide or a methacrylamide.

The term “(meth)acryloyl” group refers to an ethylenically unsaturatedgroup of formula CH₂═CR^(c)—(CO)— where R^(c) is hydrogen or methyl.

The term “N-sulfonyldicarboximide” refers to a trivalent entity offormula

The term “oxy” refers to a divalent group of formula —O—.

The term “pendant” when referring to a polymeric material means a groupthat is attached to the backbone of the polymeric material but that isnot part of the backbone of the polymeric material. The pendant group isnot involved in the polymerization reaction. For example, the group Q isthe pendant group in a polymer formed by free radical polymerization ofa reaction mixture that includes ethylenically unsaturated monomers offormula CH₂═C(R^(x))-Q. In this monomer formula, R^(x) is a hydrogen,alkyl, or aryl and Q is a group attached to the ethylenicallyunsaturated group.

The term “polymer” refers to both materials prepared from one monomersuch as a homopolymer or to materials prepared from two or more monomerssuch as a copolymer, terpolymer, or the like. Likewise, the term“polymerize” refers to the process of making a polymeric material thatcan be a homopolymer, copolymer, terpolymer, or the like.

The term “thio” refers to a divalent group of formula —S—.

The term “room temperature” refers to a temperature of about 20° C. toabout 25° C. or about 22° C. to about 25° C.

A curve connecting two groups in a formula indicates that the two groupstogether form part of a structure that can be cyclic. That is, the twogroups are linked together. A line intersecting this curve indicates acovalent bond to an atom in the structure.

The above summary of the present invention is not intended to describeeach disclosed embodiment or every implementation of the presentinvention. The description that follows more particularly exemplifiesillustrative embodiments. In several places throughout the application,guidance is provided through lists of examples, which examples can beused in various combinations. In each instance, the recited list servesonly as a representative group and should not be interpreted as anexclusive list.

DETAILED DESCRIPTION

Reaction mixtures used to prepare crosslinked polymeric materials,crosslinked polymeric materials formed from the reaction mixtures,articles containing the crosslinked polymeric materials, methods ofmaking the articles, and methods of immobilizing amine-containingmaterials are described. More specifically, the reaction mixturesinclude (a) a monomer having an amine capture group and (b) acrosslinking monomer. The crosslinked polymeric material has a pendantamine capture group that includes a N-sulfonyldicarboximide groupcapable of reacting with an amine-containing material by a ring openingreaction.

A reaction mixture is provided that includes (a) an amine capturemonomer of Formula I

and (b) a crosslinking monomer that includes at least two (meth)acryloylgroups. The monomer of Formula I can be unsubstituted or substitutedwith a halo, alkyl, alkoxy, or combinations thereof. In Formula I,

-   -   L is an oxy or —NR⁶—;    -   R¹ and R² together with a dicarboximide group to which they are        attached form a four to eight membered heterocyclic or        heterobicyclic group that can be fused to an optional aromatic        group, optional saturated or unsaturated cyclic group, or        optional saturated or unsaturated bicyclic group;    -   R³ is hydrogen or methyl;    -   R⁴ is an alkyl, aryl, aralkyl, or —N(R⁵)₂ wherein each R⁵ are        each an alkyl group or taken together with the nitrogen atom to        which they are attached form a four to eight membered        heterocyclic group;    -   R⁶ is hydrogen, alkyl, aryl, aralkyl, acyl, alkylsulfonyl, or        arylsulfonyl; and    -   Y is a single bond or a divalent group selected from an        alkylene, heteroalkylene, arylene, or combinations thereof.

The amine capture monomer of Formula I can be a (meth)acrylate monomerwhen L is oxy as shown in Formula I(a).

Some exemplary monomers are acrylates (i.e., where R³ is hydrogen) ormethacrylates (i.e., where R³ is methyl).

The amine capture monomer of Formula I can be a (meth)acrylamide monomerwhen L is —NR⁶— as shown in Formula I(b).

Some exemplary monomers are acrylamides (i.e., where R³ is hydrogen) ormethacrylamides (i.e., where R³ is methyl).

Group Y can be a single bond or can be a divalent group that includes analkylene, heteroalkylene, arylene, or combinations thereof. The divalentgroup Y can further include optional groups selected from a carbonyl,carbonylimino, carbonyloxy, oxy, thio, —NR⁶—, or combinations thereof.Group Y and the combined group —Y-L- typically do not include peroxidegroups (i.e., two oxy groups bonded together).

Group Y can be an alkylene group or Y can include an alkylene connectedto at least one other group selected from another alkylene,heteroalkylene, arylene, carbonyl, carbonyloxy, carbonylimino, oxy,thio, —NR³—, or combination thereof. In other examples, Y can be aheteroalkylene group or Y can include a heteroalkylene connected to atleast one other group selected from another heteroalkylene, alkylene,arylene, carbonyl, carbonyloxy, carbonylimino, oxy, thio, —NR⁶—, orcombination thereof. In still other examples, Y can be an arylene groupor Y can include an arylene connected to at least one other groupselected from another arylene, alkylene, heteroalkylene, carbonyl,carbonyloxy, carbonylimino, oxy, thio, —NR⁶—, or combination thereof.

In some exemplary monomers of Formula I, group Y can include acarbonyloxy, or carbonylimino group attached to an alkylene such as inthe monomer of the following formula

where n is an integer of 1 to 30. The groups R¹, R², R³, R⁴, and L arethe same as described for Formula I. In some monomers, n is no greaterthan 20, no greater than 10, no greater than 8, no greater than 6, or nogreater than 4. The monomers can be unsubstituted or substituted with ahalo, alkyl, alkoxy, or combinations thereof.

In other examples where Y includes a first alkylene group that isconnected to a second alkylene group or to a first heteroalkylene groupwith a group selected from a carbonyl, carbonyloxy, carbonylimino, oxy,thio, or —NR⁶—. Additional alkylene or heteroalkylene groups can beconnected to the second alkylene group or the first heteroalkylene groupwith a group selected from a carbonyl, carbonyloxy, carbonylimino, oxy,thio, or —NR⁶—.

Group Y can be a heteroalkylene as in the following formula

or group Y can be a heteroalkylene attached to another group such as acarboxy or carbonylimino as in the following formula

In these formulas, q is an integer of 1 to 15; x is an integer of 2 to4; and D is oxy, thio, or —NH—. The groups R¹, R², R³, R⁴, and L are thesame as previously defined for Formula I. Exemplary compounds includethose where q is an integer no greater than 12, no greater than 10, nogreater than 8, no greater than 6, no greater than 4, or no greater than2; and x is no greater than 3, or equal to 2. The compounds can beunsubstituted or substituted with a halo, alkyl, alkoxy, or combinationsthereof.

In other exemplary monomers of Formula I, group Y includes a firstheteroalkylene group that is connected to a second heteroalkylene or toa first alkylene group with a group selected from a carbonyl,carbonyloxy, carbonylimino, oxy, thio, or —NR⁶—. Additional alkylene orheteroalkylene groups can be connected to the second heteroalkylenegroup or the first alkylene group with a group selected from a carbonyl,carbonyloxy, carbonylimino, oxy, thio, or —NR⁶—.

The R⁴ group in Formula I can be an alkyl, aryl, or aralkyl. Suitablealkyl groups typically contain no greater than 30 carbon atoms, nogreater than 20 carbon atoms, no greater than 10 carbon atoms, nogreater than 6 carbon atoms, or no greater than 4 carbon atoms. In somecompounds, the alkyl group is methyl, ethyl, or propyl. Suitable arylgroups typically contain 6 to 30 carbon atoms, 6 to 24 carbon atoms, 6to 18 carbon atoms, 6 to 12 carbon atoms, or 6 carbon atoms. In somecompounds, the aryl group is phenyl. Suitable aralkyl groups typicallycontain an aryl group having 6 to 30 carbon atoms and an alkyl grouphaving no greater than 30 carbon atoms. An example of an aralkyl groupis 4-methyl-phenyl.

In other embodiments of Formula II, R⁴ is a group —N(R⁵)₂ where each R⁵are alkyl groups having no greater than 30 carbon atoms, no greater than20 carbon atoms, no greater than 10 carbon atoms, no greater than 6carbon atoms, or no greater than 4 carbon atoms. Alternatively, the twoR⁵ groups can combine together with the nitrogen atom to which they areattached to form a 4 to 8 membered ring structure. For example, the twoR⁵ groups can combine to form a five or six membered heterocyclic grouphaving a nitrogen heteroatom.

In Formula I, R¹ and R² together with a dicarboximide group to whichthey are attached form a four to eight membered heterocyclic orheterobicyclic group that can be fused to an optional aromatic group,optional saturated or unsaturated cyclic group, or optional saturated orunsaturated bicyclic group. Exemplary structures include, but are notlimited to the following:

The groups R³, R⁴, L, and Y are the same as described above for FormulaI. In these formulas, the group CH₂═CR³—CO-L-Y— can be attached to thering structure in various locations, as indicated by the placement ofthis group within the ring structure. The monomers can be unsubstitutedor substituted with a halo, alkyl, alkoxy, or combinations thereof.

Several factors can influence the selection of group Y for a particularapplication. These factors include, for example, ease of synthesis ofthe amine capture monomer, compatibility or reactivity of the aminecapture monomer with the crosslinking monomer, and reactivity orselectivity of the amine capture group with an amine-containingmaterial. For example, the size and the polarity of group Y can affectthe reactivity of the amine capture group with amine-containingmaterial. That is, varying the length and composition of group Y canalter the reactivity of the amine capture groups. Further, the size andnature of the ring structure containing the dicarboximide group canaffect the surface concentration and reactivity with theamine-containing materials. The various groups in the amine capturemonomer can be chosen, if desired, to provide a monomer that is liquidat ambient conditions. Liquid monomers tend to be useful in solventlesscoating compositions, which can be environmentally desirable.

The reaction mixtures are often coated from solution and dried (e.g.,close to 100 percent solids) prior to polymerization. Solubility of theamine capture monomer in the solvent and miscibility with thecrosslinking monomer can be important variables for obtaining suitablecoatings. These factors can be controlled by selection of group Y.Heteroalkylene groups tend to improve solubility in polar solvents andmonomers; alkylene groups tend to improve solubility in non-polarsolvents and monomers.

Exemplary amine capture monomers of Formula I include, but are notlimited to,

The amine capture monomers can be unsubstituted or substituted with ahalo, alkyl, alkoxy, or combinations thereof.

The amine capture monomers contain two reactive functional groups: anamine reactive group that includes the N-sulfonyldicarboximide group anda (meth)acryloyl group that can undergo a free-radical polymerizationreaction. Synthetic strategies must lead to these two reactivefunctional groups but be tolerant of their different reactivity. Thecompounds of Formula I may be prepared, for example, by reaction of afirst compound having a nitrogen-containing group with a second compoundcontaining a carboxylic acid anhydride. More specifically, thenitrogen-containing group of the first compound includes a nitrogen atomdirectly bonded to a sulfonyl group and to two hydrogen atoms. Thesecond compound can also include a group —Y-L-CO—CR³═CH₂. A typicalsynthesis approach is shown in Reaction Scheme A.

where R¹, R², R³, R⁴, Y, and L are the same as previously defined forFormula I.

The reaction mixture typically includes 0.1 to 90 weight percent of theamine capture monomer of Formula I based on the weight of monomers inthe reaction mixture. If less than 0.1 weight percent amine capturemonomer is included in the reaction mixture, there may not be asufficient number of pendant amine capture groups in the resultingcrosslinked polymeric material available for reaction with anamine-containing material. However, if the amount of amine capturemonomer is greater than 90 weight percent, the crosslinked polymer maynot be mechanically robust or dimensionally stable because of the lowamount of crosslinking monomer in the reaction mixture.

The reaction mixture usually contains at least 0.1 weight percent, atleast 0.5 weight percent, at least 1 weight percent, at least 2 weightpercent, at least 5 weight percent, or at least 10 weight percent of theamine capture monomer of Formula I. The reaction mixture typicallycontains up to 90 weight percent, up to 80 weight percent, up to 70weight percent, up to 60 weight percent, up to 50 weight percent, up to40 weight percent, up to 30 weight percent, or up to 20 weight percentof the amine capture monomer. For example, the reaction mixture cancontain 0.2 to 90 weight percent, 0.5 to 90 weight percent, 1 to 90weight percent, 2 to 80 weight percent, 2 to 60 weight percent, 2 to 40weight percent, 2 to 20 weight percent, or 2 to 10 weight percent of theamine capture monomer.

In addition to the monomer of Formula I, the reaction mixture contains acrosslinking monomer that includes at least two (meth)acryloyl groups.Suitable crosslinking monomers include di(meth)acrylates,tri(meth)acrylates, tetra(meth)acrylates, penta(meth)acrylates, and thelike. These (meth)acrylates can be formed, for example, by reacting(meth)acrylic acid with an alkanediols, alkanetriols, alkanetetra-ols,or alkanepenta-ols.

Exemplary crosslinking monomers include 1,2-ethanediol di(meth)acrylate;1,12-dodecanediol di(meth)acrylate; 1,4 butanediol di(meth)acrylate;1,6-hexanediol di(meth)acrylate; trimethylolpropane triacrylate (e.g.,commercially available under the trade designation TMPTA-N from SurfaceSpecialties, Smyrna, Ga. and under the trade designation SR-351 fromSartomer, Exton, Pa.); pentaerythritol triacrylate (e.g., commerciallyavailable under the trade designation SR-444 from Sartomer);tris(2-hydroxyethylisocyanurate) triacrylate (commercially availableunder the trade designation SR-368 from Sartomer); a mixture ofpentaerythritol triacrylate and pentaerythritol tetraacrylate (e.g.,commercially available from Surface Specialties under the tradedesignation PETIA with an approximately 1:1 ratio of tetraacrylate totriacrylate and under the trade designation PETA-K with an approximately3:1 ratio of tetraacrylate to triacrylate); pentaerythritoltetraacrylate (e.g., commercially available under the trade designationSR-295 from Sartomer); di-trimethylolpropane tetraacrylate (e.g.,commercially available under the trade designation SR-355 fromSartomer); ethoxylated pentaerythritol tetraacrylate (e.g., commerciallyavailable under the trade designation SR-494 from Sartomer); anddipentaerythritol pentaacrylate (e.g., commercially available under thetrade designation SR-399 from Sartomer). Mixtures of crosslinkingmonomers can be used.

The reaction mixture can contain 10 to 99.9 weight percent crosslinkingmonomer based on the weight of monomers in the reaction mixture. If lessthan this amount of crosslinking monomer is included in the reactionmixture, the resulting crosslinked polymeric material may not bemechanically robust or dimensionally stable. On the other hand, if agreater amount of crosslinking monomer is used, the number of pendantamine capture groups in the crosslinked polymeric material may be toolow to efficiently react with amine-containing materials.

The amount of crosslinking monomer can be at least 10 weight percent, atleast 15 weight percent, at least 20 weight percent, at least 30 weightpercent, at least 40 weight percent, at least 50 weight percent, atleast 60 weight percent, at least 70 weight percent, at least 80 weightpercent, or at least 90 weight percent. For example, the amount ofcrosslinking monomer can be in the range of 10 to 99 weight percent, 20to 90 weight percent, 30 to 90 weight percent, 30 to 80 weight percent,30 to 70 weight percent, or 30 to 60 weight percent.

Other optional diluent monomers such as (meth)acrylates and(meth)acrylamides can be included in the reaction mixture. Suitablediluent monomers include monomers that do not have an amine capturegroup and that have only one (meth)acryloyl group. The reaction mixturecan include up to 20 weight percent of the diluent monomer based on theweight of monomers in the reaction mixture. If more than about 20 weightpercent diluent monomer is included in the reaction mixture, thecrosslinked polymeric material may not be sufficiently mechanicallyrobust and dimensionally stable or there may not be a sufficient numberof pendant amine capture groups to effectively react withamine-containing compounds. In some reaction mixtures, no diluentmonomer is included. The amount of diluent monomer typically is in therange of 0 to 20 weight percent, 0 to 15 weight percent, 0 to 10 weightpercent, or 0 to 5 weight percent.

Some exemplary (meth)acrylate diluent monomers are alkyl (meth)acrylatessuch as methyl (meth)acrylate, ethyl (meth)acrylate, n-propyl(meth)acrylate, isopropyl (meth)acrylate, n-butyl (meth)acrylate,isobutyl (meth)acrylate, tert-butyl methacrylate, n-hexyl(meth)acrylate, cyclohexyl (meth)acrylate, 3,3,5-trimethylcyclohexyl(meth)acrylate, 2-methylbutyl (meth)acrylate, 2-ethylhexyl(meth)acrylate, 4-methyl-2-pentyl (meth)acrylate, n-octyl(meth)acrylate, isooctyl (meth)acrylate, n-decyl (meth)acrylate,isodecyl (meth)acrylate, lauryl (meth)acrylate, isononyl (meth)acrylate,isotridecyl (meth)acrylate, and behenyl (meth)acrylate.

Other exemplary (meth)acrylate diluent monomers are aryl (meth)acrylatessuch as phenyl (meth)acrylate, stearyl (meth)acrylate, and benzyl(meth)acrylate. Still other exemplary (meth)acrylate diluent monomersare hydroxy alkyl (meth)acrylates such as 2-hydroxyethyl (meth)acrylateand 3-hydroxypropyl (meth)acrylate, 4-hydroxybutyl (meth)acrylate,glycerol (meth)acrylate, and the like. Additional exemplary(meth)acrylate diluent monomers are ether-containing (meth)acrylatessuch as 2-ethoxyethyl (meth)acrylate, 2-methoxyethyl (meth)acrylate,polyethyleneglycol (meth)acrylate, and the like. The diluent monomerscan be nitrogen-containing (meth)acrylates such asN,N-dimethylaminoethyl (meth)acrylate, N,N-dimethylaminopropyl(meth)acrylate, 2-trimethylammoniumethyl (meth)acrylate halides, and thelike. The diluent monomers also can be (meth)acrylamides, N,N-dialkyl(meth)acrylamides, and the like.

Some reaction mixtures contain a solvent. Suitable solvents include, butare not limited to, water, acetonitrile, tetrahydrofuran, ethyl acetate,toluene, acetone, methyl ethyl ketone, isopropanol, N-methylpyrrolidone, chlorinated and fluorinated hydrocarbons, fluorinatedethers, or combinations thereof. Other reaction mixtures can besolventless. Exemplary solventless reaction mixtures include those inwhich the amine capture monomer is a liquid or those in which thereaction mixture is a coated composition with the solvent removed.

A crosslinked polymeric material can be prepared that includes thereaction product of the reaction mixture, as described above, thatcontains (a) an amine capture monomer of Formula I and (b) acrosslinking monomer. These reaction mixtures are typically polymerizedusing a free radical polymerization method. There is often an initiatorincluded in the reaction mixture. The initiator can be a thermalinitiator, a photoinitiator, or both. The initiator is often used at aconcentration of 0.1 to 5 weight percent, 0.1 to 3 weight percent, 0.1to 2 weight percent, or 0.1 to 1 weight percent based on the weight ofmonomers in the reaction mixture.

When a thermal initiator is added to the reaction mixture, thecrosslinked polymer can be formed at room temperature (i.e., 20 to 25degrees Celsius) or at an elevated temperature. The temperature neededfor polymerization often depends on the particular thermal initiatorused. Examples of thermal initiators include organic peroxides or azocompounds.

When a photoinitiator is added to the reaction mixture, a crosslinkedpolymeric material can be formed by the application of actinic radiationuntil the composition gels or hardens. Suitable actinic radiationincludes electromagnetic radiation in the infrared region, visibleregion, ultraviolet region, or a combination thereof.

Examples of photoinitiators suitable in the ultraviolet region include,but are not limited to, benzoin, benzoin alkyl ethers (e.g., benzoinmethyl ether and substituted benzoin alkyl ethers such anisoin methylether), phenones (e.g., substituted acetophenones such as2,2-dimethoxy-2-phenylacetophenone and substituted alpha-ketols such as2-methyl-2-hydroxypropiophenone), phosphine oxides, polymericphotoinitiators, and the like.

Commercially available photoinitiators include, but are not limited to,2-hydroxy-2-methyl-1-phenyl-propane-1-one (e.g., commercially availableunder the trade designation DAROCUR 1173 from Ciba Specialty Chemicals),a mixture of 2,4,6-trimethylbenzoyl-diphenyl-phosphine oxide and2-hydroxy-2-methyl-1-phenyl-propan-1-one (e.g., commercially availableunder the trade designation DAROCUR 4265 from Ciba Specialty Chemicals),2,2-dimethoxy-1,2-diphenylethan-1-one (e.g., commercially availableunder the trade designation IRGACURE 651 from Ciba Specialty Chemicals),a mixture of bis(2,6-dimethoxybenzoyl)-2,4,4-trimethyl-pentylphosphineoxide and 1-hydroxy-cyclohexyl-phenyl-ketone (e.g., commerciallyavailable under the trade designation IRGACURE 1800 from Ciba SpecialtyChemicals), a mixture ofbis(2,6-dimethoxybenzoyl)-2,4,4-trimethyl-pentylphosphine oxide (e.g.,commercially available under the trade designation IRGACURE 1700 fromCiba Specialty Chemicals),2-methyl-1[4-(methylthio)phenyl]-2-morpholinopropan-1-one (e.g.,commercially available under the trade designation IRGACURE 907 fromCiba Specialty Chemicals), 1-hydroxy-cyclohexyl-phenyl-ketone (e.g.,commercially available under the trade designation IRGACURE 184 fromCiba Specialty Chemicals),2-benzyl-2-(dimethylamino)-1-[4-(4-morpholinyl)phenyl]-1-butanone (e.g.,commercially available under the trade designation IRGACURE 369 fromCiba Specialty Chemicals), bis(2,4,6-trimethylbenzoyl)-phenylphosphineoxide (e.g., commercially available under the trade designation IRGACURE819 from Ciba Specialty Chemicals), ethyl 2,4,6-trimethylbenzoyldiphenylphosphinate (e.g., commercially available from BASF, Charlotte, N.C.under the trade designation LUCIRIN TPO-L), and2,4,6-trimethylbenzoyidiphenylphosphine oxide (e.g., commerciallyavailable from BASF, Charlotte, N.C. under the trade designation LUCIRINTPO).

Photoinitiators suitable for use in the visible region often include anelectron donor, and electron acceptor such as an iodonium salt, and avisible light sensitizing compound such as an alpha di-ketone. Suchphotoinitiators are further described, for example, in U.S. PatentPubiications 2005/0113477 A1 (Oxman et al.) and 2005/0070627 A1 (Falsafiet al.); and U.S. Pat. No. 6,765,036 B2 (Dede et al.), all incorporatedherein by reference.

An article can be prepared that includes a substrate and a crosslinkedpolymeric material disposed on a surface of the substrate. Thecrosslinked polymeric material is often formed after coating a surfaceof the substrate with a reaction mixture that contains (a) an aminecontaining monomer of Formula I and (b) a crosslinking monomer that hasat least two (meth)acryloyl groups.

The substrates can have any useful form including, but not limited to,films, sheets, membranes, filters, nonwoven or woven fibers, hollow orsolid beads, bottles, plates, tubes, rods, pipes, or wafers. Thesubstrates can be porous or non-porous, rigid or flexible, transparentor opaque, clear or colored, and reflective or non-reflective. Thesubstrates can have a flat or relatively flat surface or can have atextured surface such as wells, indentations, channels, bumps, or thelike. The substrates can have a single layer or multiple layers ofmaterial. Suitable substrate materials include, for example, polymericmaterials, glasses, ceramics, metals, metal oxides, hydrated metaloxides, or combinations thereof.

Suitable polymeric substrate materials include, but are not limited to,polyolefins (e.g., polyethylene and polypropylene), polystyrenes,polyacrylates, polymethacrylates, polyacrylonitriles, polyvinylacetates, polyvinyl alcohols, polyvinyl chlorides, polyoxymethylenes,polycarbonates, polyamides, polyimides, polyurethanes, phenolics,polyamines, amino-epoxy resins, polyesters, silicones, cellulose basedpolymers, polysaccharides, or combinations thereof.

Suitable glass and ceramic substrate materials can include, for example,silicon, aluminum, lead, boron, phosphorous, zirconium, magnesium,calcium, arsenic, gallium, titanium, copper, or combinations thereof.Glasses typically include various types of silicate containingmaterials.

In some embodiments, the substrate includes a layer of diamond-likeglass as disclosed in U.S. Pat. No. 6,696,157 B1 (David et al.), thedisclosure of which is incorporated herein by reference in its entirety.The diamond-like glass is an amorphous material that includes carbon,silicon, and one or more elements selected from hydrogen, oxygen,fluorine, sulfur, titanium, or copper. Some diamond-like glass materialsare formed from a tetramethysilane precursor using a plasma process. Ahydrophobic material can be produced that is further treated in anoxygen plasma to control the silanol concentration on the surface.

Suitable metals, metal oxides, or hydrated metal oxides for substratescan contain, for example, gold, silver, platinum, palladium, aluminum,copper, chromium, iron, cobalt, nickel, zinc, and the like. Themetal-containing material can be alloys such as stainless steel, indiumtin oxide, and the like. In some embodiments, a metal-containingmaterial is the top layer of a multilayer substrate. For example, thesubstrate can have a polymeric second layer and a metal containing firstlayer. In one example, the second layer is a polymeric film and thefirst layer is a thin film of gold. In other examples, a multilayersubstrate includes a polymeric film coated with a titanium-containinglayer and then coated with a gold-containing layer. That is, thetitanium layer can function as a tie layer or a primer layer foradhering the layer of gold to the polymeric film. In other examples of amultilayer substrate, a silicon support layer is covered with a layer ofchromium and then with a layer of gold. The chromium layer can improvethe adhesion of the gold layer to the silicon layer.

The articles can be prepared by providing a substrate, disposing areaction mixture on a surface of the substrate, and curing the reactionmixture to form a crosslinked polymeric material. The reaction mixturecontains (a) an amine capture monomer of Formula I and (b) acrosslinking monomer, as described above. The reaction mixture can becured, for example, through the application of heat (i.e., a thermalinitiator is used) or through the application of actinic radiation(i.e., a photoinitiator is used).

The reaction mixture can often wet the surface of the substrate and theresulting crosslinked polymer adheres to the surface of the substrate.The crosslinked polymer usually adheres without the formation of acovalent bond between a reactive group on the polymer and acomplementary group on the surface of the substrate. Rather, the polymeradheres by interlocking with surface imperfections on the substrate.

In some articles, the crosslinked polymeric material is patterned on thesubstrate. Any suitable pattern of the crosslinking polymeric materialcan be formed. For example, the pattern can be in the form of text,design, image, or the like. The pattern can be, for example, in the formof dots, squares, rectangles, circles, lines, or waves (e.g., squarewaves, sinusoidal waves, and saw tooth waves).

One method of forming a patterned crosslinked polymeric layer includesdisposing a pattern of the reaction mixture on the surface of thesubstrate by screen printing, jet printing (e.g., spray jet, valve jet,or ink jet printing), and the like. Useful devices forjet printing aredescribed, for example, in U.S. Pat. No. 6,513,897 (Tokie) and in U.S.Patent Publication No. 2002/0128340 (Young et al.), both incorporatedherein by reference. After application of the pattern to the surface ofthe substrate, the reaction mixture can be cured. For example, thereaction mixture can be cured by application of heat if a thermalinitiator is included in the reaction mixture or by application ofactinic radiation if a photoinitiator is included in the reactionmixture.

Another method of forming a patterned crosslinked polymeric layerincludes forming a layer of the reaction mixture on the substrate,curing a first portion of the reaction mixture to form a pattern ofcrosslinked polymeric material on the substrate, and removing a secondportion of the reaction mixture that is not cured. The layer of reactionmixture on the substrate can be prepared using any suitable techniquesuch as, for example, brush coating, spray coating, gravure coating,transfer roll coating, knife coating, curtain coating, wire coating, anddoctor blade coating.

One method of polymerizing a portion of the reaction mixture involvesthe use of a photoinitiator in the reaction mixture and the use ofmasks. The mask contains a pattern of openings and can be positionedbetween the layer of reaction mixture and the actinic radiation source.Actinic radiation can pass through the openings in the mask. Uponexposure to actinic radiation, a first portion of the reaction mixturelayer corresponding to the openings in the mask can polymerize and asecond portion of the reaction mixture layer that is blocked from theactinic radiation by the mask remains uncured or not reacted. That is,the uncured reaction mixture is a monomeric composition that has notgelled or hardened to form a crosslinked polymeric material. The uncuredreaction mixture can be removed using a suitable solvent for themonomers of Formula I, the crosslinking monomers, and any optionaldiluent monomers. The solvent typically does not dissolve the curedcrosslinked polymeric material because of the extensive crosslinking.Thus, the uncured reactive mixture can be removed leaving a pattern ofcrosslinked polymeric material on the substrate surface.

Suitable solvents for removing the unreacted reaction mixture include,but are not limited to, water, acetonitrile, tetrahydrofuran, ethylacetate, toluene, acetone, methyl ethyl ketone, isopropanol, N-methylpyrrolidone, chlorinated and fluorinated hydrocarbons, fluorinatedethers, or combinations thereof. The crosslinked polymeric material istypically insoluble in these solvents. As used herein, an insolublepolymer is one that has a solubility at room temperature that is lessthan 0.01 weight percent in a solvent such as, for example, water,acetonitrile, tetrahydrofuran, ethyl acetate, toluene, acetone, methylethyl ketone, isopropanol, N-methyl pyrrolidone, chlorinated andfluorinated hydrocarbons, fluorinated ethers, or combinations thereof.

Suitable masks for this method of patterning include polymeric materials(e.g., polyesters such as polyethylene terephthalate or polyethylenenaphthalate, polyimide, polycarbonate, or polystyrene), metal foilmaterials (e.g., stainless steel, other steels, aluminum, or copper),paper, woven or nonwoven fabric materials, or combinations thereof.Polymeric masks and the openings in these masks are further described inU.S. Pat. No. 6,897,164 B2 (Baude et al.), incorporated herein byreference. The openings in the mask can be of any suitable dimension.

The crosslinked polymeric material has pendant amine capture groups.Thus, an article having a layer or pattern of crosslinked polymericmaterial on a surface of a substrate can have amine capture groups thatare capable of reacting with an amine-containing material. The pendantamine capture groups of the polymeric material are of Formula II

attached to the backbone of the crosslinked polymeric backbone. Thegroups L, Y, R¹, R² and R⁴ are previously described for Formula I. Theasterisk in Formula II indicates the location where the pendant aminecapture group is attached to the backbone of the crosslinked polymericmaterial. The pendant amine capture groups include aN-sulfonyldicarboximide group.

The pendant groups according to Formula II usually have improvedhydrolytic stability compared to derivatives of N-hydroxysuccinimide,which are groups known to react with amine-containing materials. Becauseof the improved hydrolytic stability of the pendant groups of FormulaII, the crosslinked polymeric materials and articles containing thecrosslinked polymeric material typically can be used in aqueous systems.

The amine capture groups can be reacted with an amine-containingmaterial resulting in the immobilization of the amine-containingmaterial. A method of immobilizing an amine-containing material includesproviding a substrate, and disposing a reaction mixture on a surface ofthe substrate. The reaction mixture contains (a) a monomer of Formula Iand (b) a crosslinking monomer that includes at least two (meth)acryloylgroups. The method further includes curing the reaction mixture to forma crosslinked polymeric material having a pendant amine capture group,and reacting an amine-containing material with the pendant amine capturegroup of the crosslinked polymeric material. In this method, theN-sulfonyldicarboximide group in the pendant amine capture group canreact with an amine-containing material. In some embodiments, theamine-containing materials are biological materials such as, forexample, amino acid, peptide, DNA, RNA, protein, enzyme, organelle,cells, tissue, immunoglobin, or a fragment thereof. In otherembodiments, the amine-containing material is a non-biological aminesuch as an amine-containing analyte.

The amine-containing material can react with the pendant amine capturegroup by a ring opening reaction with the N-sulfonyldicarboximide group.The amine-containing material can have a primary amine group or asecondary amine group. For example, the amine-containing material can bea primary amine-containing biological material of formula H₂N-T where Tis the remainder of the primary amine-containing biological material(i.e., the group T is equal to a primary amine-containing biologicalmaterial of formula H₂N-T minus the —NH₂ group). Group T often has analkylene group attached directly to the —NH₂ group. The primaryamine-containing biological material reacts with the crosslinkedpolymeric material having pendant groups of Formula II to produce acrosslinked polymeric material having a reacted pendant group of FormulaIII:

In Formula III, R⁷ is an alkylene having 1 to 5 carbon atoms, aromaticgroup, saturated or unsaturated cyclic group, saturated or unsaturatedbicyclic group, or combination thereof. R⁷ corresponds to the combinedgroups R¹ and R² in Formulas I and II. The group L, Y, R⁴, and * are thesame as previously defined for Formulas I and II. The presence of theimmobilized amine can be determined, for example, using massspectroscopy, contact angle measurement, infrared spectroscopy, andellipsometry. Additionally, various immunoassays and optical microscopictechniques can be used if the amine-containing material is abiologically active material.

The rate of reaction of amine-containing materials with theN-sulfonyldicarboximide groups of the pendant amine capture groups ofFormula II is typically faster than the rate of hydrolysis of theN-sulfonyidicarboximide group. That is, immobilization ofamine-containing materials occurs at a faster rate than the hydrolysisreactions. The amine-containing materials are not easily displaced onceimmobilization to a pendant group of the crosslinked polymeric materialhas occurred due to the formation of a covalent carbonylimino bond.

Immobilized biological amine-containing materials can be useful in themedical diagnosis of a disease or of a genetic defect. The immobilizedamine-containing materials can also be used for biological separationsor for detection of the presence of various biomolecules. Additionally,the immobilized amine-containing materials can be used in bioreactors oras biocatalysts to prepare other materials. The pendant groups with theN-sulfonylaminocarbonyl groups also can be used to detectamine-containing analytes that are not biological materials. Theamine-containing analytes can have primary amine groups, secondary aminegroups, or a combination thereof.

Other materials can be further bound to the immobilized amine-containingmaterial. This further bound material can be associated with theamine-containing material before immobilization of the amine-containingmaterial or can be bound to the amine-containing material subsequent toimmobilization of the amine-containing material. The amine-containingmaterial and the further bound material can be complementary RNAfragments, complementary DNA fragments, an antigen-antibody combination,an immunoglobin-bacterium combination, and the like.

Biological amine-containing materials often can remain active afterattachment to the pendant group of the crosslinked polymeric material(i.e., the pendant groups according to Formula III can includebiologically active amine-containing materials). For example, animmobilized antibody can subsequently bind to an antigen or animmobilized antigen can subsequently bind to an antibody. Similarly animmobilized amine-containing biological material that has a portion thatcan bind to a bacterium can subsequently bind to the bacterium (e.g., animmobilized immunoglobulin can subsequently bind to a bacterium such asStaphylococcus aureus).

The pendant groups of Formula II and Formula III are attached to thebackbone of a crosslinked polymeric material. The crosslinked polymericmaterial is formed by a free radical polymerization reaction of(meth)acryloyl groups. A crosslinked polymeric material with pendantgroups of Formula II can be further crosslinked by reaction of a primaryamine containing material having at least two primary amine groups,secondary amine groups, or a combination thereof. That is, an aminecontaining material that has more than one primary amine or secondaryamine groups may react with more than one pendant group of Formula II.

EXAMPLES

These examples are merely for illustrative purposes only and are notmeant to be limiting on the scope of the appended claims. All parts,percentages, ratios, etc. in the examples and the rest of thespecification are by weight, unless noted otherwise. Solvents and otherreagents used were obtained from Sigma-Aldrich Chemical Company;Milwaukee, Wis. unless otherwise noted.

Table of Abbreviations Abbreviation or Trade Designation Description ACNAcetonitrile DMF Dimethylformamide NMP N-methylpyrrolidinone THFTetrahydrofuran TMPTA Trimethylolpropane triacrylate PhotoinitiatorDAROCUR 1173 Photo curing agent 2-hydroxy-2-methyl-1-phenyl-1-propanone,available from Ciba; Hawthorne, NJ DI water Deionized water PEIPolyethyleneimine IPA Isopropyl alcohol TWEEN-25 Polyoxyethylenesorbitanmonolaurate from Sigma, St Louis, MO

Preparative Example 1 Preparation of

A 3-neck round bottom flask, fitted with a magnetic stir bar, anaddition funnel and a hose adapter that was connected to a source ofnitrogen gas, was charged with a 60 weight percent dispersion of NaH inmineral oil (9.45 g) and hexane (20 mL). The mixture was stirred forapproximately 15 minutes after which DMF (100 mL) was added to theflask. A mixture of p-toluenesulfonamide (15.7 g) and5-norbornene-2,3-dicarboxylic anhydride (16.2 g) in DMF (100 mL) wasslowly added to the flask from the addition funnel. The mixture wasallowed to stir overnight at room temperature. An additional solution of5-norbornene-2,3-dicarboxylic anhydride (1.6 g) in DMF (10 mL) was addeddropwise to the flask and the mixture was stirred for approximately 6hours. Acetic anhydride (28.14 g) was then added to the flask and themixture was stirred overnight. Aqueous NaHCO₃ solution was then added tothe flask, followed by aqueous HCl. The mixture was filtered and thefiltered solid was dried overnight using a vacuum oven. The solid wasthen recrystallized from methanol to afford 14.8 g of product.

Preparative Example 2 Preparation of

A mixture of 2-mercaptoethanol (1.69 grams), theN-(4-methylbenzenesulfonyl)succinimide product of Preparative Example 1(7.0 grams), VAZO 64 (0.2 grams), and ethyl acetate (8.7 grams) waspurged with nitrogen gas for approximately 15 minutes and was thenheated at 55° C. under a nitrogen atmosphere for 24 hours. The solutionwas allowed to cool to room temperature, during which time whitecrystals precipitated. The crystals were isolated by filtration, washedwith ethyl acetate and dried to afford 3.2 grams of product.

Preparative Example 3 Preparation of

To a magnetically stirred mixture of the alcohol product of PreparativeExample 2 (5.0 grams), NMP (20 milliliters), and triethylamine (1.5grams) within a round bottom flask there is slowly added acryloylchloride (1.25 grams) over about 15 minutes. The mixture is stirredovernight at room temperature under a nitrogen atmosphere, after whichtime the mixture is poured into 0.1N aqueous HCl (100 milliliters) in abeaker. This mixture is extracted with ethyl acetate and then the ethylacetate mixture is dried over sodium sulfate. The volatile componentsare then removed using a rotary evaporator to afford the product.

Preparative Example 4 Preparation of

A solution of the N-(4-methylbenzenesulfonyl)dicarboximide product ofPreparative Example 1 (25.0 g) and mercaptoacetic acid (7.63 g) in ethylacetate (161 g) within a 3-neck round bottom flask was purged withnitrogen gas for 10 minutes. The flask was fitted with a refluxcondenser and a magnetic stir bar. Azobis(isobutyronitrile) (0.0484 g)was added to the flask and the mixture was stirred and heated at 55° C.for sixteen hours. After the mixture had cooled to room temperature, thesolvent was removed using a rotary evaporator. The brown solid wasrecrystallized from toluene/acetonitrile to give 11.9 g of product.

Preparative Example 5 Preparation of

A mixture of the carboxy-containing product of Preparative Example 4(5.06 g), thionyl chloride (1.62 g), DMF (1 drop), and chloroform (61.8g) was made within a 100 mL round bottom flask. The flask was fittedwith a magnetic stir bar and a reflux condenser and then the mixture wasstirred and heated under reflux for two hours. The mixture was allowedto cool to room temperature and then heptane was added to the flaskuntil the product precipitated. The mixture was filtered and the productwas dried in a vacuum oven to give 4.8 g of product.

Preparative Example 6 Preparation of

To a round bottom flask fitted with a magnetic stirrer was added ethylacetate (0.25 milliliter), 2-hydroxyethyl methacrylate (0.10 gram) andN-ethyldiisopropylamine (0.12 gram). The product of Preparative Example5 (0.30 gram) was dissolved in ethyl acetate (1.75 milliliter) and addedto the flask under a nitrogen atmosphere and left to stir at roomtemperature overnight. After which time was added ethyl acetate (10milliliter). The mixture was poured into 0.1N aqueous HCl (10milliliter) in a beaker. The aqueous layer was separated and the organicphase was washed with 0.1N aqueous HCl (10 milliliter), followed by asaturated sodium chloride solution (10 milliliter). The ethyl acetatemixture was dried over magnesium sulfate. A rotary evaporator was usedto concentrate the mixture, which was purified on an Analogix FlashChromatography system over 20 minutes with a gradient from 35:63 to85:15 of ethyl acetate and hexanes. The fractions containing the desiredproduct were collected and the volatile components were then removedusing a rotary evaporator to afford 0.2178 gram of product, a yield of59%.

Preparative Example 7 Preparation of

To a magnetically stirred mixture of the alcohol product of PreparativeExample 2 (5.0 grams), NMP (20 milliliters), and triethylamine (1.5grams) within a round bottom flask there is slowly added methacrylicanhydride (2.12 grams) over about 15 minutes. The mixture is stirredovernight at room temperature under a nitrogen atmosphere, after whichtime the mixture is poured into 0.1N aqueous HCl (100 milliliters) in abeaker. This mixture is extracted with ethyl acetate and then the ethylacetate mixture is dried over sodium sulfate. The volatile componentsare then removed using a rotary evaporator to afford the product.

Preparative Example 8 Preparation of

A mixture of methanesulfonamide (10 grams), 1,2,4-benzenetricarboxylicanhydride (26.3 grams), triethylamine (37.2 grams), and DMF (84.6 grams)was magnetically stirred under a nitrogen atmosphere overnight at roomtemperature. The mixture was then heated to 50° C. and stirred at thistemperature for 30 minutes, after which time it was allowed to cool toroom temperature and was filtered. The solid was recrystallized fromglacial acetic acid and the white crystalline solid was filtered, washedwith diethyl ether, and dried to afford 5.4 grams of product.

Preparative Example 9 Preparation of

A mixture of the carboxylic acid product of Preparative Example 8, (3.0grams), thionyl chloride (1.72 grams), DMF (1 drop), and ACN (18.9grams) within a round bottom flask fitted with a reflux condenser wasmagnetically stirred under a nitrogen atmosphere at reflux for 1 hour.The volatile components were then removed using a rotary evaporator andthe partially solid residue in the flask was washed into a fritted glassfunnel with diethyl ether. The solid was washed with diethyl ether andwas dried under a flow of nitrogen gas to afford 2.9 grams of product.

Preparative Example 10 Preparation of

A mixture of the carbonyl chloride product of Preparative Example 9 (5.0grams), NMP (20 milliliters), 2-hydroxyethyl acrylate (1.97 grams), andtriethylamine (1.89 grams) within a round bottom flask is magneticallystirred overnight under a nitrogen atmosphere at room temperature. Themixture is then poured into 0.1N aqueous HCl (100 milliliters) in abeaker. This mixture is extracted with ethyl acetate and then the ethylacetate mixture is dried over sodium sulfate. The volatile componentsare then removed using a rotary evaporator to afford the product.

Preparative Example 11 Preparation of

A mixture of the carbonyl chloride product of Preparative Example 9(1.06 grams), acetonitrile (5.5 milliliters), 2-hydroxyethylmethacrylate (0.51 grams), and ethyidiisopropylamine (0.62 grams) withina round bottom flask was magnetically stirred overnight under a nitrogenatmosphere at room temperature. The mixture was concentrated on a rotaryevaporator. The resulting oil was taken up in chloroform (25milliliters) then washed with 0.1N HCl (25 milliliters) and dried overmagnesium sulfate. The chloroform was removed resulting in an oil thatwas taken up in hot methanol. Upon cooling a dark oil settled and theclear solution was decanted, concentrated on a rotary evaporator. Theresulting oil was dissolved in hot isopropyl alcohol, upon cooling toroom temperature a white crystalline solid precipitated, which wasfiltered, washed with chilled IPA and dried in an oven to provide 81milligrams of desired product.

Preparative Example 12 Preparation of

A mixture of the carbonyl chloride product of Preparative Example 9 (5.0grams), NMP (20 milliliters) N-phenylethanolamine (2.34 grams), andtriethylamine (1.89 grams) within a round bottom flask is magneticallystirred overnight under a nitrogen atmosphere at room temperature. Themixture is then poured into 0.1N aqueous HCl (100 milliliters) in abeaker. This mixture is extracted with ethyl acetate and then the ethylacetate mixture is dried over sodium sulfate. The volatile componentsare then removed using a rotary evaporator to afford the product.

Preparative Example 13 Preparation of

A mixture of the alcohol product of Preparative Example 12 (5.0 grams),NMP (20 milliliters), acryloyl chloride (1.18 grams), and triethylamine(1.45 grams) within a round bottom flask is magnetically stirredovernight under a nitrogen atmosphere at room temperature. The mixtureis then poured into 0.1N aqueous HCl (100 milliliters) in a beaker. Thismixture is extracted with ethyl acetate and then the ethyl acetatemixture is dried over sodium sulfate. The volatile components are thenremoved using a rotary evaporator to afford the product.

Preparative Example 14 Preparation of

A mixture of the alcohol product of Preparative Example 12 (5.0 grams),NMP (20 milliliters), methacrylic anhydride (2.00 grams), andtriethylamine (1.45 grams) within a round bottom flask is magneticallystirred overnight under a nitrogen atmosphere at room temperature. Themixture is then poured into 0.1N aqueous HCl (100 milliliters) in abeaker. This mixture is extracted with ethyl acetate and then the ethylacetate mixture is dried over sodium sulfate. The volatile componentsare then removed using a rotary evaporator to afford the product.

Preparative Example 15 Preparation of:

To a round bottom flask fitted with a magnetic stirrer was added ethylacetate (0.25 mL), 2-hydroxyethyl methacrylate (0.10 g) andN-ethyldiisopropylamine (0.12 g). The product of Preparative Example 5(0.30 g) was dissolved in ethyl acetate (1.75 mL) and added to the flaskunder a nitrogen atmosphere and left to stir ambient overnight. Afterwhich time was added ethyl acetate (10 mL). The mixture was poured into0.1N aqueous HCl (10 mL) in a beaker. The aqueous layer was separatedand the organic phase was washed with 0.1N aqueous HCl (10 mL), followedby a saturated sodium chloride solution (10 mL). The ethyl acetatemixture was dried over magnesium sulfate. A rotary evaporator was usedto concentrate the mixture, which was purified on an Analogix FlashChromatography system over 20 minutes with a gradient from 35:63 to85:15 of ethyl acetate and hexanes. The fractions containing the desiredproduct were collected and the volatile components were then removedusing a rotary evaporator to afford 0.2178 g of product, a yield of 59%.

Preparation of PEI Coated Glass Slide Matrix

Glass microscope slides were soaked for two hours in a 5 Molar NaOHbath; rinsed with DI water, ethanol, and methanol; and dried under astream of nitrogen. The clean slides were kept in an 80° C. oven untilneeded.

Eight of the slides were arranged in a matrix of 2 columns and 4 rows.The slides were secured in position within the matrix using a strip oftape down the middle of the back side of each slide. A 2 weight percentsolution of PEI in IPA was coated onto the matrix using a number 12Mayer rod. The coating was allowed to dry in air.

Preparation of IgG Labeled with Cy5

The contents of three vials of Cy5 dye (3H-Indolium,2-[5-[1-[6-[(2,5-dioxo-1-pyrrolidinyl)oxy]-6-oxohexyl]-1,3-dihydro-3,3-dimethyl-5-sulfo-2H-indol-2-ylidene]-1,3-pentadienyl]-1-ethyl-3,3-dimethyl-5-sulfo-,inner salt (9CI)) were dissolved in dimethylsulfoxide (DMSO) to a totalvolume of 100 microliters. The vials of Cy5 dye were obtained fromGE-Amersham Biosciences, Piscataway N.J. The resulting dye solution wasadded to 1 milliliter of a 5 milligrams/milliliter solution of mouse IgGin 0.1 M sodium carbonate (at pH 9.0). The mouse IgG was obtained fromSigma, St. Louis, Mo. The resulting solution was protected from lightexposure and gently rocked for 45 minutes at room temperature. Thissolution contained Cy5-labelled antibody and unreacted Cy5.

Cy5-labelled antibody (Cy5-IgG) was separated from unreacted Cy5 labelusing gel filtration chromatography. The solution containing the Cy5-IgGand unreacted CyS was added to a PD-10 column that was equilibratedusing phosphate buffer solution (PBS) at pH 7.4. The PD-10 column wasobtained from GE-Amersham Biosciences, Piscataway N.J. The Cy5-IgGfraction was collected by washing with PBS at pH 7.4. The Cy5/ IgG ratiowas calculated by measuring the IgG concentration (280 nm) and the Cy5concentration (650 nm) in the Cy5-IgG fraction. The productspecifications provided by the manufacturer of Cy5 and IgG were followedto obtain the extinction coefficient values for IgG and Cy5 as well asthe absorbance contribution from covalently bound Cy5 at 280 nm. Thefinal CyS-IgG solution had a concentration 1.3 milligrams/milliliterCy5-IgG with a Cy5/IgG ratio of 2.2. From this stock solution, testsolutions containing 130, 50, 13 and 5 micrograms/milliliters of Cy5-IgGin 10 millimolar carbonate buffer at pH 9.6 were prepared. The bufferwas prepared using sodium carbonate/sodium bicarbonate buffer capsulesavailable from Sigma, St. Louis, Mo.

Comparative Example C1

A solution of Photoinitiator (0.01 gram) and TMPTA (0.6 gram) in ACN (5grams) was prepared. This solution contained 11 weight percent TMPTA and0.18 weight percent Photoinitiator based on the weight of the solution.This solution was coated onto a PEI coated glass slide matrix, which isdescribed above, using a number 12 Mayer rod. The coating was allowed todry in air. The coated matrix was passed twice through a curingapparatus at a rate of 50 feet/minute (15 meters/minute). The curingapparatus was obtained from Fusion Systems of Gaithersburg, Md. and wasequipped with a F300 lamp. The resulting cured coating was rubbed with ametal spatula to ensure the coating could not be rubbed off.

Spots of 5 microliters of the Cy5-IgG test solutions prepared above anda sample without Cy5-IgG were applied to the coated surface and allowedto sit for 30 minutes. The surface was rinsed with 0.25 weight percentTWEEN-25 in DI water and then rinsed with DI water. The slides weredried under nitrogen and placed into a fluorescence reader, which iscommercially available from Tecan Group LTD, Research Triangle Park,N.C. under the trade designation LS SERIES TECAN. Single scanmeasurements were made by adjusting the focal height to 1002micrometers, 40 micrometers resolution, oversampling of 3 micrometers, again of 160, and a pinhole depth focus of ±150 micrometers. The data wasanalyzed as 16-bit pixelized TIFF files using software commerciallyavailable from Molecular Devices Corp, Sunnyvale, Calif. under the tradedesignation GENEPIX PRO. The data are shown in Table 1.

Example 1

A solution was prepared by dissolving the material from PreparativeExample 6 (0.031 gram), Photoinitiator (0.009 gram), and TMPTA (0.54gram) in ACN (5.2 grams). The resulting solution contained 0.5 weightpercent Preparative Example 1, 0.18 weight percent Photoinitiator, and10 weight percent TMPTA based on the weight of the solution. Thissolution was coated onto a PEI coated glass slide matrix describedabove, using a number 12 Mayer rod. The resulting coating was allowed todry in air. The coated matrix was passed twice at a rate of 50feet/minute (15 meters/minute) through the Fusion Systems curingapparatus equipped with a F300 lamp. The resulting cured coating wasrubbed with a metal spatula to ensure the coating could not be rubbedoff.

Five microliter spots of the Cy5-IgG test solutions and of a samplewithout Cy5-IgG were made onto the coated surface and allowed to sit for30 minutes. The surface was rinsed with 0.25 weight percent TWEEN-25 inDI water and then with DI water. The slides were dried under nitrogenand placed into a fluorescence reader, which is commercially availablefrom Tecan Group LTD, Research Triangle Park, N.C. under the tradedesignation LS SERIES TECAN. Single scan measurements were made byadjusting the focal height to 1002 micrometers and using 40 micrometersresolution, a gain of 160, oversampling of 3 micrometers, and a pinholedepth focus of ±150 micrometers. The data was analyzed as 16-bitpixelized TIFF files using software commercially available fromMolecular Devices Corp, Sunnyvale, Calif. under the trade designationGENEPIX PRO. The data are shown in Table 1.

Example 2

A solution was prepared by dissolving the material of PreparativeExample 11 (0.032 gram), Photoinitiator (0.009 gram), and TMPTA (0.56gram) in ACN (5.3 grams). The resulting solution contained 0.5 weightpercent Preparative Example 2, 0.18 weight percent photoinitiator, and10 weight percent TMPTA based on the weight of the solution. Thissolution was coated onto a PEI coated glass slide matrix described aboveusing a number 12 Mayer rod. The resulting coating was allowed to dry inair.

The coated matrix was passed twice through the Fusion Systems curingapparatus equipped with a F300 lamp at a rate of 50 feet/minute (15meters/minute). The resulting cured coating was rubbed with a metalspatula to ensure the coating could not be rubbed off.

Five microliter spots of the Cy5-IgG test solutions and of a samplewithout Cy5-IgG were applied to the coated surface and allowed to sitfor 30 minutes. The surface was rinsed with 0.25 weight percent TWEEN-25in DI water and then with DI water. The slides were dried under nitrogenand placed into a fluorescence reader, which is commercially availablefrom Tecan Group LTD, Research Triangle Park, N.C. under the tradedesignation LS SERIES TECAN. Single scan measurements were made byadjusting the focal height to 1002 micrometers and using 40 micrometersresolution, a gain of 160, oversampling of 3 micrometers, and a pinholedepth focus of ±150 micrometers. The data was analyzed as 16-bitpixelized TIFF files using software commercially available fromMolecular Devices Corp, Sunnyvale, Calif. under the trade designationGENEPIX PRO. The data are shown in Table 1.

TABLE 1 Concentration of Fluorescence Fluorescence Fluorescence Cy5-IgGMeasurement Measurement Measurement (Micrograms/milliliter) Example C1Example 1 Example 2 130 34108 65535 65535 50 17295 58371 36596 13 813542565 13915 5 1270 18540 4079 0 1266 288 1400 *Note the value of 65535is the maximum number of pixels that can be measured using a 16-bitfluorescence reader.

The complete disclosures of the patents, patent documents, andpublications cited herein are incorporated by reference in theirentirety as if each were individually incorporated. Variousmodifications and alterations to this invention will become apparent tothose skilled in the art without departing from the scope and spirit ofthis invention. It should be understood that this invention is notintended to be unduly limited by the illustrative embodiments andexamples set forth herein and that such examples and embodiments arepresented by way of example only with the scope of the inventionintended to be limited only by the claims set forth herein as follows.

1. A reaction mixture comprising: a) an amine capture monomer of FormulaI

wherein L is an oxy or —NR⁶—; R¹ and R² together with a dicarboximidegroup to which they are attached form a four to eight memberedheterocyclic or heterobicyclic group that can be fused to an optionalaromatic group, optional saturated or unsaturated cyclic group, oroptional saturated or unsaturated bicyclic group; R³ is hydrogen ormethyl; R⁴ is an alkyl, aryl, aralkyl, or —N(R⁵)₂ wherein each R⁵ is analkyl group or both R⁵ groups taken together with the nitrogen atom towhich they are attached form a four to eight membered heterocyclicgroup; R⁶ is hydrogen, alkyl, aryl, aralkyl, acyl, alkylsulfonyl, orarylsulfonyl; Y is a single bond or a divalent group comprising analkylene, heteroalkylene, arylene, or combinations thereof; the aminecapture monomer is unsubstituted or substituted with a halo, alkyl,alkoxy, or combinations thereof; and b) a crosslinking monomercomprising at least two (meth)acryloyl groups.
 2. The reaction mixtureof claim 1, wherein the amine capture monomer is of Formula I(a)


3. The reaction mixture of claim 1, wherein the amine capture monomer isof Formula I(b)


4. The reaction mixture of claim 1, wherein the amine capture monomer isof formula

wherein n is an integer of 1 to
 30. 5. The reaction mixture of claim 1,wherein the amine capture monomer is of formula

wherein q is an integer of 1 to 15; x is an integer of 2 to 4; and D isoxy, thio, or —NH—.
 6. The reaction mixture of claim 1, wherein theamine capture monomer is of formula


7. The reaction mixture of claim 1, wherein the amine capture monomercomprises


8. The reaction mixture of claim 1, further comprising a photoinitiator,a thermal initiator, or a combination thereof.
 9. The reaction mixtureof claim 1, wherein Y further comprises a carbonyl, carbonyloxy,carbonylimino, oxy, thio, —NR⁶—, or combinations thereof.
 10. Thereaction mixture of claim 1, wherein the reaction mixture comprises 0.1to 50 weight percent of the amine capture monomer of Formula I based onthe weight of the monomers in the reaction mixture.